Immersion type microscope objective

ABSTRACT

An immersion type microscope objective is configured by, in order from the object side to an image side, a positive lens group Ga including a cemented lens obtained by cementing a plano-convex lens whose plane surface faces the object side to a meniscus lens whose concave surface faces the object side, and a positive single lens, a positive lens group Gb including a cemented lens, a lens group Gc including at least one cemented lens, a lens group Gd having a meniscus lens having a strongly concave surface that faces the image side, and a lens group Ge having a negative lens having a strongly concave surface that faces the object side.

CROSS REFERENCE TO RELATED APPLICATIONS

This application claims benefit of Japanese Applications No.2006-333338, filed Dec. 11, 2006, and No. 2007-311963, filed Dec. 3,2007, the contents of which are incorporated by this reference.

BACKGROUND OF THE INVENTION

1. Field of the Invention

The present invention relates to the technology of a microscopeobjective and a fluorescent observation apparatus.

2. Description of the Related Art

Conventionally, fluorescent observation apparatuses are used inobserving the behavior of molecules inside biological specimen withutilizing fluorescent tags. By connecting fluorescence with specificbiological molecules, fluorescent tags can be used in observing thebehavior, the coupling status, the moving status, etc. of molecules in abiological specimen. A fluorescent tag can be a fluorescent dye, afluorescent protein, a quantum dot, etc.

One of the recent trends is observation with multicolored fluorescenttags. With the multicolored fluorescent tags, plural types of moleculescan be discriminated in an observation, and complicated events such asan interaction of molecules in a living specimen.

Using the multicolored fluorescent tags, a wider range of wavelength isrequired in the fluorescent observation than in conventionalobservations.

In addition, wavelengths of 440 nm and 405 nm are used forphotoactivation fluorescent protein such as Kaede, PA-GFP, etc. Suchshort wavelength light has not been frequently used in the conventionalapplications.

As a result, there is an increasing request for an objectiveaberration-corrected in a wider wavelength range than the conventionalobjective.

The target of the observation in a living specimen was the structure ofthe cells, but the target recently is tending toward the behavior ofmolecules. As a result, the resolution in the focal plane and theoptical axis of microscopes is getting higher request. In addition,since confocal microscopes are popular, an observation can be performedwith higher resolution. That is, there has also been request for anobjective of high numerical aperture to attain higher resolution.

The Japanese Published Patent Application No. H7-35983 and the JapanesePublished Patent Application No. 2003-21786 disclose microscopeobjectives with high numerical aperture appropriate for fluorescentobservations.

SUMMARY OF THE INVENTION

The immersion type microscope objective as one of the embodiments of thepresent invention is configured by, in order from the object side to theimage side, a positive lens group Ga, a positive lens group Gb, a lensgroup Gc, a lens group Gd, and a lens group Ge. The positive lens groupGa includes a cemented lens obtained by cementing a plano-convex lenswhose plane surface faces the object side to a meniscus lens whoseconcave surface faces the object side, and a positive single lens. Thepositive lens group Gb is formed by a cemented lens. The lens group Gcincludes one or more cemented lens. The lens group Gd includes ameniscus lens having a strongly concave surface that faces the imageside. The lens group Ge includes a negative lens having a stronglyconcave surface that faces the object side. Assuming that H1 indicatesthe ray height of the marginal ray emergent from the lens group Gb, H2indicates the ray height of the marginal ray incident to the lens groupGd, f indicates the focal length of the entire objective system, andf(Gb) indicates the focal length of the lens group Gb, the followingconditions (1) and (2) are satisfied.0.5≦H2/H1≦0.75.  (1)7.8≦|f(Gb)/f|≦20.  (2)

The fluorescent observation apparatus as one of the embodiments of thepresent invention includes: a light source; a beam splitting deviceselectively reflecting light from the light source; an objective forilluminating or observing a specimen; a stage for fixing the specimen; awavelength selection device for selecting a desired wavelength rangebased on rays that pass through the beam splitting device; and adetector for detecting light that passes through the wavelengthselection device. With the configuration, assume that λ1 is the longestwavelength and λ2 is the shortest wavelength in illuminating thespecimen using two or more wavelengths, and Δ1 and Δ2 are the focalpoint of λ1 and λ2 respectively on the optical axis of the objective.Based on the assumption, the following conditions are satisfied.λ1−λ2≧180 nm.  (11)|Δ1−Δ2|<0.2 μm.  (12)

BRIEF DESCRIPTION OF THE DRAWINGS

The present invention will be more apparent from the following detaileddescription when the accompanying drawings are referenced.

FIG. 1 is a sectional view of the objective according to the embodiment1;

FIG. 2 shows the aberration of the objective according to the embodiment1;

FIG. 3 is a sectional view of the objective according to the embodiment2;

FIG. 4 shows the aberration of the objective according to the embodiment2;

FIG. 5 is a sectional view of the objective according to the embodiment3;

FIG. 6 shows the aberration of the objective according to the embodiment3;

FIG. 7 is a sectional view of the objective according to the embodiment4;

FIG. 8 shows the aberration of the objective according to the embodiment4;

FIG. 9 is a sectional view of the objective according to the embodiment5;

FIG. 10 shows the aberration of the objective according to theembodiment 5;

FIG. 11 is a sectional view of the objective according to the embodiment6;

FIG. 12 shows the aberration of the objective according to theembodiment 6;

FIG. 13 is a sectional view of the objective according to the embodiment7;

FIG. 14 shows the aberration of the objective according to theembodiment 7;

FIG. 15 is a sectional view of the objective according to the comparisonexample 1;

FIG. 16 shows the aberration of the objective according to thecomparison example 1;

FIG. 17 is a sectional view of the objective according to the comparisonexample 2;

FIG. 18 shows the aberration of the objective according to thecomparison example 2;

FIG. 19 is a sectional view of the objective according to the comparisonexample 3;

FIG. 20 shows the aberration of the objective according to thecomparison example 3;

FIG. 21 is a sectional view of the tube lens A;

FIG. 22 is a sectional view of the tube lens B;

FIG. 23 shows the focal point on the axis at each wavelength in theobject side according to each embodiment;

FIG. 24A shows the outline of the view of the aberration of the axialchromatic aberration of the objective in conventional art;

FIG. 24B shows the outline of the view of the aberration of the axialchromatic aberration of the objective in embodiments of the presentinvention;

FIG. 25 shows the outline of the difference in the focal point of eachwavelength depending on the difference in level of the illuminationluminous flux;

FIG. 26 shows the chromatic aberration of magnification at field number9 on each wavelength according to each embodiment;

FIG. 27 shows the laser scanning confocal microscope in conventionalart;

FIG. 28 shows the laser scanning type mirror focal microscope accordingto an embodiment of the present invention;

FIG. 29 shows the image processing system using the laser scanning typemirror focal microscope according to an embodiment of the presentinvention; and

FIGS. 30A, 30B, and 30C show an example of the observation of thefluorescent resonant energy transfer (FRET) according to the presentinvention.

DESCRIPTION OF THE PREFERRED EMBODIMENTS

The immersion type microscope objective as one of the embodiments of thepresent invention is configured by, in order from the object side to theimage side, a positive lens group Ga, a positive lens group Gb, a lensgroup Gc, a lens group Gd, and a lens group Ge. The positive lens groupGa includes a cemented lens obtained by cementing a plano-convex lenswhose plane surface faces the object side to a meniscus lens whoseconcave surface faces the object side, and a positive single lens. Thepositive lens group Gb is formed by a cemented lens. The lens group Gcincludes one or more cemented lens. The lens group Gd includes ameniscus lens having a strongly concave surface that faces the imageside. The lens group Ge includes a negative lens having a stronglyconcave surface that faces the object side. Assuming that H1 indicatesthe ray height of the marginal ray emergent from the lens group Gb, H2indicates the ray height of the marginal ray incident to the lens groupGd, f indicates the focal length of the entire objective, and f(Gb)indicates the focal length of the lens group Gb, the followingconditions (1) and (2) are satisfied.0.5≦H2/H1≦0.75.  (1)7.8≦|f(Gb)/f|≦20.  (2)

The condition (1) indicates the condition of appropriately correctingthe axial chromatic aberration. Based on the condition (1), the rayheight from the lens group Gb to the lens group Gd is regulated, and theaxial chromatic aberration can be easily corrected. When the H2/H1 islower than the left hand side of the condition (1), the ray height fromthe lens group Gb to the lens group Gd is too low, and it is difficultto correct the distortion or the comatic aberration. If the H2/H1 ishigher than the right hand side of the condition (1), the ray heightfrom the lens group Gb to the lens group Gd is too high, and it is hardto appropriately correct the axial chromatic aberration.

The condition (2) indicates the condition of appropriately correctingvarious aberrations such as a spherical aberration etc. Based on thecondition (2), the power of the lens group Gb is regulated. If the|f(Gb)/f| is lower than the left hand side of the condition (2), thepower of the lens group Gb is too strong, and the amount of generatedspherical aberration is too large. If the |f(Gb)/f| is higher than theright hand side of the condition (2), then the power of the lens groupGb is too weak, it is difficult to satisfy the conditional expression(1), and the axial chromatic aberration cannot be appropriatelycorrected.

More advantageously, the following conditions (1)′ and (2)′ issatisfied.0.55≦H2/H1≦0.73.  (1)′8≦|f(Gb)/f|≦20.  (2)′

The lens group Ge includes a negative lens Len whose concave surfacefaces the object side, and a positive lens Lep. If νd(Len) is defined asthe Abbe number of the glass of the negative lens Len, and νd(Lep) isdefined as the Abbe number of the glass of the positive lens Lep, thenit is desired that the following conditional expression (3) issatisfied.60≧νd(Len)−νd(Lep)≧25.  (3)

The condition (3) is to correct the chromatic aberration of themagnification by setting a large difference in Abbe number between thenegative lens and the positive lens of the lens group Ge. If the lefthand side of the condition is lower than 25, the chromatic aberration ofthe magnification cannot be sufficiently corrected. Thecompensation-free objective for correcting the axial chromaticaberration and the chromatic aberration of magnification using only anobjective is allowed the flexibility in selecting combination units(image optics system and illumination optics system). Moreadvantageously, the following conditions (3)′ is satisfied.45≧νd(Len)−νd(Lep)≧30.  (3)′

The optical glass of the positive lens Lep is an optical glasscontaining ingredient Nb₂O₅ or Ta₂O₅. When the refractive index of thed-line of the positive lens Lep is defined as nd(Lep), it is desiredthat the following conditions are satisfied.1.65≦nd(Lep)≦1.8  (4)25≦νd(Lep)≦41  (5)

To appropriately correct the comatic aberration and the chromaticaberration of the magnification, it is necessary that the positive lensLep is made of an optical glass having a high refractive index and largechromatic dispersion (low Abbe number νd). The optical glassescontaining ingredient Nb₂O₅ or Ta₂O₅ have a high refractive index andchromatic dispersion. Therefore, the comatic aberration and thechromatic aberration of the magnification can be appropriatelycorrected. Furthermore, these optical glasses have low autofluorescenceand high transparency in the ultraviolet range. Therefore, it ispossible to make an observation with high contrast and brightness in thefluorescent observation.

It is also desired that the lens group Gb is a cemented triplet of apositive lens, a negative lens, and a positive lens. The axial chromaticaberration can be appropriately corrected by arranging the cementedtriplet in the lens group Gb having a high ray height of a marginal ray.

Furthermore, it is also desired to satisfy the following condition (6)when H3 is defined as the ray height of the marginal ray emergent fromthe lens group Ge.0.5≦H3/H1≦0.65  (6)

The condition is set to appropriately correct various aberrations. Ifthe H3/H1 is lower than the left hand side of the condition, then thepower of the lens group Ga and the lens group Gb is too weak, strongpower is required as the subsequent group to the lens group Gc, and theamount of generated aberrations becomes high. If the H3/H1 is higherthan the right hand side of the condition, then the power of the lensgroup Ga and lens group Gb is too strong, and the amount of generatedaberrations in the lens group Ga and the lens group Gb becomes large.

In addition, the positive lens group Ga includes: a cemented lens G1obtained by cementing a plano-convex lens whose plane surface faces theobject side to a meniscus lens whose concave surface faces the objectside; a positive meniscus lens G2 whose concave surface faces the objectside; and a positive single lens G3.

Assuming that f(G1+G2) indicates the composite focal length of thecemented lens G1 and the meniscus lens G2, f indicates the focal lengthof the entire objective, and D indicates the length of the marginal raybetween the cemented lens G1 and the meniscus lens G2, the configurationsatisfying the following conditional expressions (7) and (8) is alsodesired.1≦f(G1+G2)/f≦2  (7)D/f≦0.6  (8)

To collect the dispersed luminous flux from the object, it is commonthat the lens group on the object side has strong power. Specifically,with the immersion type objective having high NA, the lens group closestto the object generally has the geometrically close to a hemisphere.However, if the lens group closest to the object is assigned highconvergence power, the spherical aberration or the axial chromaticaberration generated there are too large and it is difficult toappropriately correct them. According to the present invention, thestrong convergence effect assigned only to the lens group closest to theobject is also assigned to the two lens groups, that is, the cementedlens G1 and the meniscus lens G2, thereby reducing the amount ofgenerated aberration, and increasing the lens surface for which theaberration is to be corrected.

The conditions (7) and (8) regulate the lens groups G1 and G2. If thef(G1+G2) is lower than left hand side of the condition (7), then thepower of the lens groups G1 and G2 is too strong, and the amount of thespherical aberration and the amount of the axial chromatic aberrationincrease. If the f(G1+G2) is higher than the right hand side of thecondition (7), then no sufficient convergence power can be obtained, theray height in the subsequent lens groups becomes higher, and it isdifficult to appropriately correct the spherical aberration and theaxial chromatic aberration. If the D/f is higher than right hand side ofthe condition (8), the balance of the power between the lens groups G1and G2 cannot be maintained and the aim to replace the conventional lensgroup with two lens groups cannot be attained. By satisfying theconditional expressions (7) and (8), the lens group G1 is alike to theshape of a hemisphere, and the lens group G2 also alike to the shape ofa hemisphere. That is, there are two sagittally deep shape lenses. Withthe configuration, the load of the refractive power of the lens group G1is also shared by the lens group G2, and the effect of the commondifference of the thickness of the lens of the lens group G1 can bedispersed to the lens group G2, thereby improving the workability of thelens group G1.

Furthermore, the characteristic of the appearance of the objective as anembodiment of the present invention is that the radius of a curvature onthe image side of the cemented lens G1 is less than the focal length,and the radius of a curvature on the image side of the meniscus lens G2is less than two times the focal length f. Based on the characteristics,the ray height of and subsequent to the lens group G3 can be reduced, ahigh order aberration (spherical aberration, comatic aberration) can beeasily corrected, and the outer diameter of the entire lens can bereduced, thereby improving the workability of the lens.

It is desired that the lens group Gc can be moved in the direction ofthe optical axis, and the following conditional expression (9) can besatisfied.|f(Gc)/f|≦50  (9)

Here, f indicates the focal length of the entire system, and f(Gc)indicates the focal length of the lens group Gc.

When an observation is made deeply in the cell of a living specimen(refractive index of 1.33 through 1.45), the generation of the sphericalaberration caused by the difference in refractive index can be moreefficiently suppressed by a smaller difference in refractive indexbetween a cell and oil. However, if the refractive index of the oil isless than 1.51, the refractive index difference from the cover glass(nd=1.52426) is large. Therefore, when the thickness of the cover glassis shifted from 0.17 mm, a large spherical aberration occurs. Thecondition (9) is to correct the spherical aberration from the thicknessof the cover glass and the spherical aberration from the refractiveindex difference between the cells of a living body (nd=1.33 through1.45) and the oil. When the |f(Gc)/f| is higher than the right hand sideof the condition (9), the spherical aberration cannot be sufficientlycorrected.

It is also considered and desired that the following condition (10) canbe satisfied in place of the conditional expression (9).10≦|f(Gc)/f|≦20  (10)

If the |f(Gc)/f| is lower than 10, then the power of the lens group Gcis too strong, and the amount of generated above-mentioned in the lensgroup Gc increases. Furthermore, the amount of movement of the lensgroup Gc decreases, and the operability is degraded when a correctionlink for correcting the spherical aberration is used.

Next, the embodiment of the microscope objective according to thepresent invention is described below with reference to each embodiment.

In the embodiments, r is the radius of a curvature (in mm) of each lenssurface, d is the interval (in mm) between lens surfaces, nd is therefractive index of the d-line of each lens, νd is the Abbe number inthe d-line of each lens. β is the magnification, NA is the numericalaperture, WD is the operation distance (in mm), f is the focal length(in mm) of the entire system of an objective, f(G1+G2) is the compositefocal length (in mm) of the first lens group and the second lens group,f(Ga) through f(Ge) are the focal lengths (in mm) of each lens group ofthe lens group Ga through the lens group Ge, H1 is the ray height (inmm) of the marginal rays emergent from the lens group Gb, H2 is the rayheight (in mm) of the marginal rays incident to the lens group Gd, andH3 is the ray height (in mm) of the marginal rays on the emission sideof the objective.

The optical glass used in each of the following embodiments describedbelow is excellent in transparency in an ultraviolet range, an opticalglass with less autofluorescence is selected, and each embodiment showsthe optimum objective for a fluorescent observation. In addition, eachoptical glass is an environment-friendly glass (nonlead glass), and eachembodiment is an objective in consideration for the environment.

The refractive index of each wavelength of each medium is describedbelow.

h-LINE g-LINE F-LINE e-LINE d-LINE C-LINE WAVELENGTH (nm) 404.656435.835 486.13 546.07 587.56 656.27 νd COVER 1.54076 1.53629 1.530981.52656 1.52426 1.52133 54.3 GLASS OIL 1 1.53735 1.53105 1.52373 1.517811.51483 1.51116 41.0 OIL 2 1.41772 1.41409 1.40975 1.40615 1.404301.40197 52.0 GLASS 1.52972 1.52617 1.52187 1.51822 1.51630 1.51383 64.21 GLASS 1.92090 1.91048 1.89820 1.88813 1.88298 1.87654 40.8 2 GLASS1.60531 1.60099 1.59579 1.59139 1.58909 1.58615 61.1 3 GLASS 1.582281.57869 1.57435 1.57070 1.56879 1.56638 71.4 4 GLASS 1.50712 1.504431.50115 1.49837 1.49692 1.49506 81.5 5 GLASS 1.66407 1.65693 1.648501.64154 1.63797 1.63345 42.4 6 GLASS 1.44641 1.44439 1.44192 1.439831.43873 1.43731 95.2 7 GLASS 1.49892 1.49591 1.49224 1.48911 1.487451.48531 70.3 8 GLASS 1.77954 1.76771 1.75420 1.74341 1.73800 1.7313032.2 9 GLASS 1.46968 1.46675 1.46318 1.46014 1.45852 1.45642 67.8 10GLASS 1.61847 1.61436 1.60938 1.60518 1.60298 1.60016 65.4 11 GLASS1.63723 1.63071 1.62300 1.61664 1.61336 1.60922 44.5 12 GLASS 1.634511.63010 1.62479 1.62033 1.61800 1.61504 63.3 13 GLASS 1.84951 1.835171.81904 1.80633 1.80000 1.79224 29.8 14 GLASS 1.79917 1.79197 1.783371.77621 1.77250 1.76780 49.6 15

Each embodiment is an infinity correction type objective in which therays emergent from the objective are parallel luminous flux, and doesnot form an image by itself. For example, the following tube lens A(focal length of 180 mm), or tube lens B (focal length of 180 mm) isused with embodiments. Here, FIG. 21 is a sectional view of the tubelens A, and FIG. 22 is a sectional view of the tube lens B.

(tube lens A) surface r d nd νd 1 68.7541 7.7321 1.48749 70.23 2−37.5679 3.4742 1.80610 40.92 3 −102.8477 0.6973 4 84.3099 6.02381.83400 37.16 5 −50.7100 3.0298 1.64450 40.82 6 40.6619

(tube lens B) surface r d nd νd 1 214.5976 5.7000 1.60300 65.44 2−52.2905 3.8192 1.51633 64.14 3 152.7590 17.9239 4 101.1841 8.29661.48749 70.23 5 −54.0793 3.8834 1.61340 44.27 6 −288.2268

When the tube lens A is used in combination, the interval between theobjective in each embodiment and the tube lens A can be in the rangefrom 50 mm to 170 mm. When the tube lens B is used in combination, theinterval between the objective in each embodiment and the tube lens Bcan be in the range from 50 mm to 250 mm. The aberration in eachembodiment shown below is obtained by combining with the tube lens A atthe interval of 120 mm.

Here, the aberration diagrams are expressed in the objective plane whichcomputed by the ray tracing in the direction from the image plane(photoreceptive plane) to the object plane (plane of the specimen),which include a spherical aberration, offence against the sinecondition, a astigmatism, and a comatic aberration. From the aberrationdiagrams, the objective according to an embodiment of the presentinvention is appropriately corrected on the aberration such as the axialchromatic aberration, chromatic aberration of magnification, etc. from405 nm to 656 nm.

Embodiment 1

The embodiment 1 according to the present invention has theconfiguration shown in FIG. 1. That is, it is configured by, in orderfrom the object side to the image side, a positive lens group Ga, apositive lens group Gb, a lens group Gc, a lens group Gd, and a lensgroup Ge. The positive lens group Ga includes a cemented lens G1including a plano-convex lens whose plane surface faces the object sideand a meniscus lens whose concave surface faces the object side, and apositive single lens G2. The positive lens group Gb is formed by acemented triplet G3 including a positive lens, a negative lens, and apositive lens. The lens group Gc is formed by a cemented lens G4including a negative lens and a positive lens, and a cemented lens G5including a positive lens and a negative lens. The lens group Gd isformed by a cemented lens G6 including a positive lens and a negativelens, and shaped a meniscus lens which has the strongly concave surfacethat faces the image side. The lens group Ge is formed by a negativelens G7 and a positive lens G8.

The lens data according to the embodiment 1 is listed below. Theobjective according to the embodiment 1 is used with the immersingsolution of nd=1.51483, νd=41.0. The immersing solution is conventionaloil for a microscope objective.

TABLE 1 surface r d nd νd Medium  1 INF 0.1700 1.52426 54.3 COVER GLASS 2 INF 0.1600 1.51483 41.0 OIL 1  3 INF 0.5300 1.51630 64.2 GLASS 1  4−1.2824 3.6211 1.88298 40.8 GLASS 2  5 −3.1856 0.1200  6 −30.9403 4.47921.56879 71.4 GLASS 4  7 −7.3998 0.1983  8 17.1268 6.4359 1.49692 81.5GLASS 5  9 −8.9524 1.0000 1.61336 44.5 GLASS 12 10 15.0118 5.33641.43873 95.2 GLASS 7 11 −12.8352 0.1998 12 120.6880 1.0000 1.63797 42.4GLASS 6 13 9.8562 5.1029 1.43873 95.2 GLASS 7 14 −21.6730 0.2000 1515.4221 3.6001 1.43873 95.2 GLASS 7 16 −15.5126 1.0000 1.63797 42.4GLASS 6 17 34.3563 0.2000 18 7.4348 4.6350 1.60298 65.4 GLASS 11 19−23.8545 2.7618 1.63797 42.4 GLASS 6 20 4.4583 2.3996 21 −3.9504 0.80001.61800 63.3 GLASS 13 22 306.4907 2.3235 23 −19.2670 3.0954 1.80000 29.8GLASS 14 24 −7.7632 β = 60x, NA = 1.4, field number = 22, WD = 0.16, f =3 f (Ga) = 4.809 f (Gb) = 27.656 f (Gc) = 156.491 f (Gd) = −97.432 f(Ge) = −32.359 f (Ga) = 1.60 f (Gb) = 9.22 f (Gc) = 52.16 f (Gd) =−32.48 f (Ge) = −10.79 H1 = 7.27 H2 = 5.2 H3 = 4.2 νd (Len) = 63.3 νd(Lep) = 29.8 nd (Len) = 1.61800 nd (Lep) = 1.80000 (1) H2/H1 = 0.72 (2)f (Gb)/f = 9.22 (3) νd (Len) − νd (Lep) = 33.5 (6) H3/H1 = 0.58

With the above-mentioned configuration, the aberration correction on thechromatic aberration and various aberrations are corrected in thewavelength range from 405 nm to 656 nm according to the presentembodiment. The aberration correction is shown in FIG. 2.

Embodiment 2

The embodiment 2 according to the present invention has theconfiguration shown in FIG. 3. That is, it is configured by, in orderfrom the object side to the image side, a positive lens group Ga, apositive lens group Gb, a lens group Gc, a lens group Gd, and a lensgroup Ge. The positive lens group Ga includes a cemented lens G1including a plano-convex lens whose plane surface faces the object and ameniscus lens whose concave surface faces the object side, and apositive single lens G2. The positive lens group Gb is formed by acemented triplet G3 including a positive lens, a negative lens, and apositive lens. The lens group Gc is formed by a cemented lens G4including a negative lens and a positive lens, and a cemented lens G5including a positive lens and a negative lens. The lens group Gd isformed by a cemented lens G6 including a positive lens and a negativelens, and shaped a meniscus lens which has the strongly concave surfacethat faces the image side. The lens group Ge is formed by a negativelens G7 and a positive lens G8.

The lens data according to the embodiment 2 is listed below. Theobjective according to the embodiment 2 is used with the immersingsolution of nd=1.51483, νd=41.0. The immersing solution is conventionaloil for a microscope objective.

TABLE 2 surface r d nd νd Medium  1 INF 0.1700 1.52426 54.3 COVER GLASS 2 INF 0.1600 1.51483 41.0 OIL 1  3 INF 0.5300 1.51630 64.2 GLASS 1  4−1.3252 3.8164 1.88298 40.8 GLASS 2  5 −3.2544 0.1200  6 77.8515 4.71601.49692 81.5 GLASS 5  7 −8.4959 0.1983  8 16.9794 6.8055 1.49692 81.5GLASS 5  9 −8.5793 1.0000 1.61336 44.5 GLASS 12 10 18.1665 4.19181.43873 95.2 GLASS 7 11 −13.7643 0.2004 12 58.7464 1.0000 1.63797 42.4GLASS 6 13 9.2538 3.9412 1.43873 95.2 GLASS 7 14 −29.3071 0.2000 1513.0273 3.4902 1.43873 95.2 GLASS 7 16 −13.4308 1.0000 1.63797 42.4GLASS 6 17 30.3754 0.1996 18 6.7126 4.6876 1.60298 65.4 GLASS 11 19−9.0871 1.7678 1.63797 42.4 GLASS 6 20 4.0409 2.4000 21 −3.5269 0.80001.48745 70.3 GLASS 8 22 INF 4.2261 23 −18.8036 2.9073 1.80000 29.8 GLASS14 24 −8.8041 β = 60x, NA = 1.4, field number = 22, WD = 0.16, f = 3 f(Ga) = 4.587 f (Gb) = 28.014 f (Gc) = 158.735 f (Gd) = −68.519 f (Ge) =−36.452 f (Ga) = 1.53 f (Gb) = 9.34 f (Gc) = 52.91 f (Gd) = −22.84 f(Ge) = −12.15 H1 = 6.91 H2 = 4.72 H3 = 4.2 νd (Len) = 70.3 νd (Lep) =29.8 nd (Len) = 1.48745 nd (Lep) = 1.80000 (1) H2/H1 = 0.68 (2) f (Gb)/f= 9.34 (3) νd (Len) − νd (Lep) = 40.5 (6) H3/H1 = 0.61 (9) |f (Gc)/f| =52.9

With the above-mentioned configuration, the aberration correction on thechromatic aberration and various aberrations are corrected in thewavelength range from 405 nm to 656 nm according to the presentembodiment. The aberration correction is shown in FIG. 4.

Embodiment 3

The embodiment 3 according to the present invention has theconfiguration shown in FIG. 5. That is, it is configured by, in orderfrom the object side to the image side, a positive lens group Ga, apositive lens group Gb, a lens group Gc, a lens group Gd, and a lensgroup Ge. The positive lens group Ga includes a cemented lens G1including a plano-convex lens whose plane surface faces the object sideand a meniscus lens whose concave surface faces the object side, apositive single lens G2, and a positive single lens G3. The positivelens group Gb is formed by a cemented triplet G4 including a positivelens, a negative lens, and a positive lens. The lens group Gc is formedby a cemented lens G5 including a negative lens and a positive lens, anda cemented lens G6 including a positive lens and a negative lens. Thelens group Gd is formed by a cemented lens G7 including a positive lensand a negative lens, and shaped a meniscus lens which has the stronglyconcave surface that faces the image side. The lens group Ge is formedby a cemented lens G8 including a negative lens and a positive lens. Thelens data according to the embodiment 3 is listed below. The objectiveaccording to the embodiment 3 is used with the immersing solution ofnd=1.51483, νd=41.0. The immersing solution is conventional oil for amicroscope objective.

TABLE 3 surface r d nd νd Medium  1 INF 0.1700 1.52426 54.3 COVER GLASS 2 INF 0.1600 1.51483 41.0 OIL 1  3 INF 0.4800 1.51630 64.2 GLASS 1  4−1.3140 3.5370 1.88298 40.8 GLASS 2  5 −3.0604 0.1496  6 INF 4.49431.43873 95.2 GLASS 7  7 −7.3891 0.1498  8 30.0757 3.1123 1.43873 95.2GLASS 7  9 −16.1821 0.1495 10 INF 4.4336 1.49692 81.5 GLASS 5 11 −8.01291.0000 1.63797 42.4 GLASS 6 12 15.9529 4.5207 1.49692 81.5 GLASS 5 13−12.1334 0.3151 14 20.1446 3.4835 1.43873 95.2 GLASS 7 15 −12.43071.0000 1.63797 42.4 GLASS 6 16 26.5927 0.1497 17 14.8592 3.1218 1.4969281.5 GLASS 5 18 −11.8346 1.0000 1.63797 42.4 GLASS 6 19 246.5698 0.149720 5.1806 4.3972 1.49692 81.5 GLASS 5 21 −15.1849 1.0823 1.77250 49.6GLASS 1 22 3.4182 2.6989 23 −3.3601 0.7917 1.48745 70.3 GLASS 8 24−21.5242 6.3630 1.73800 32.2 GLASS 9 25 −7.6321 β = 60x, NA = 1.4, fieldnumber = 22, WD = 0.16, f = 3 f (Ga) = 4.057 f (Gb) = 55.132 f (Gc) =424.929 f (Gd) = −14.706 f (Ge) = 425.459 f (Ga) = 1.35 f (Gb) = 18.38 f(Gc) = 141.64 f (Gd) = −4.90 f (Ge) = 141.82 H1 = 6.7 H2 = 4.19 H3 = 4.2νd (Len) = 70.3 νd (Lep) = 32.2 nd (Len) = 1.48745 nd (Lep) = 1.73800(1) H2/H1 = 0.63 (2) f (Gb)/f = 18.38 (3) νd (Len) − νd (Lep) = 38.1 (6)H3/H1 = 0.63 (9) |f (Gc)/f| = 141.6

With the above-mentioned configuration, the aberration correction on thechromatic aberration and various aberrations are corrected in thewavelength range from 405 nm to 656 nm according to the presentembodiment. The aberration correction is shown in FIG. 6.

Embodiment 4

The embodiment 4 according to the present invention has theconfiguration shown in FIG. 7. That is, it is configured by, in orderfrom the object side to the image side, a positive lens group Ga, apositive lens group Gb, a lens group Gc, a lens group Gd, and a lensgroup Ge. The positive lens group Ga includes a cemented lens G1including a plano-convex lens whose plane surface faces the object sideand a meniscus lens whose concave surface faces the object side, apositive single lens G2, and a positive single lens G3. The positivelens group Gb is formed by a cemented triplet G4 including a positivelens, a negative lens, and a positive lens. The lens group Gc is formedby a cemented lens G5 including a negative lens and a positive lens, anda cemented lens G6 including a positive lens and a negative lens. Thelens group Gd is formed by a cemented lens G7 including a positive lensand a negative lens, and shaped a meniscus lens which has the stronglyconcave surface that faces the image side. The lens group Ge is formedby a cemented lens G8 including a negative lens and a positive lens. Thelens data according to the embodiment 4 is listed below. The objectiveaccording to the embodiment 4 is used with the immersing solution ofnd=1.51483, νd=41.0. The immersing solution is conventional oil for amicroscope objective.

TABLE 4 surface r d nd νd Medium  1 INF 0.1700 1.52426 54.3 COVER GLASS 2 INF 0.1598 1.51483 41.0 OIL 1  3 INF 0.4800 1.51630 64.2 GLASS 1  4−1.3953 3.6786 1.88298 40.8 GLASS 2  5 −3.1587 0.1484  6 INF 4.41691.43873 95.2 GLASS 7  7 −7.5017 0.1497  8 46.9496 2.4748 1.43873 95.2GLASS 7  9 −21.4389 0.1609 10 64.4978 4.9058 1.49692 81.5 GLASS 5 11−8.4012 1.0000 1.63797 42.4 GLASS 6 12 25.5835 3.9330 1.49692 81.5 GLASS5 13 −12.8982 0.1496 14 14.3371 2.3534 1.43873 95.2 GLASS 7 15 −89.13991.0000 1.63797 42.4 GLASS 7 16 7.6109 0.5000 17 7.1829 5.1309 1.4969281.5 GLASS 5 18 −9.1388 1.0000 1.63797 42.4 GLASS 6 19 −118.5923 0.149920 5.2788 4.2673 1.49692 81.5 GLASS 5 21 −9.2055 1.0823 1.77250 49.6GLASS 15 22 3.5809 2.6995 23 −3.3555 1.0007 1.48745 70.3 GLASS 8 24−22.6626 6.1219 1.73800 32.2 GLASS 9 25 −7.6084 β = 60x, NA = 1.4, fieldnumber = 22, WD = 0.16, f = 3 f (Ga) = 4.178 f (Gb) = 39.814 f (Gc) =133.751 f (Gd) = −13.142 f (Ge) = 350.726 f (Ga) = 1.39 f (Gb) = 13.27 f(Gc) = 44.58 f (Gd) = −4.38 f (Ge) = 116.91 H1 = 6.96 H2 = 4.1 H3 = 4.2νd (Len) = 70.3 νd (Lep) = 32.2 nd (Len) = 1.48745 nd (Lep) = 1.73800(1) H2/H1 = 0.59 (2) f (Gb)/f = 13.27 (3) νd (Len) − νd (Lep) = 38.1 (6)H3/H1 = 0.60

With the above-mentioned configuration, the aberration correction on thechromatic aberration and various aberrations are corrected in thewavelength range from 405 nm to 656 nm according to the presentembodiment. The aberration correction is shown in FIG. 8.

Embodiment 5

The embodiment 5 according to the present invention has theconfiguration shown in FIG. 9. That is, it is configured by, in orderfrom the object side to the image side, a positive lens group Ga, apositive lens group Gb, a lens group Gc, a lens group Gd, and a lensgroup Ge. The positive lens group Ga includes a cemented lens G1including a plano-convex lens whose plane surface faces the object sideand a meniscus lens whose concave surface faces the object side, and apositive single lens G2. The positive lens group Gb is formed by acemented triplet G3 including a positive lens, a negative lens, and apositive lens. The lens group Gc is formed by a cemented lens G4including a negative lens and a positive lens, and a cemented lens G5including a positive lens and a negative lens. The lens group Gd isformed by a cemented lens G6 including a positive lens and a negativelens, and shaped a meniscus lens which has the strongly concave surfacethat faces the image side. The lens group Ge is formed by a negativelens G7 and a positive lens G8. The lens group Gc can move in thedirection of the optical axis.

The data according to the embodiment 5 is listed below. The objectiveaccording to the embodiment 5 is used with the immersion liquid ofnd=1.40430, νd=52.0. The immersion liquid can be silicone oil. Therefractive index of the silicone oil is close to the refractive index ofthe cells of a living specimen (nd=1.33 through 1.45), and there is fewrefractive index mismatch between the silicone oil and the cells of aliving specimen. Therefore, the objective using silicone oil has theadvantage that the cells of a living body can be clearly and deeplyobserved. In addition, it has less autofluorescence, and is appropriatefor a fluorescent observation. Furthermore, since it is not volatile andnot easily changed in refractive index, it is appropriate for along-time observation. On the other hand, a commonly used glycerinimmersing solution and an immersing solution as a mixture of glycerinand water are inappropriate for a long-time observation because theabsorbent glycerin changes its refractive index with the lapse of time.

TABLE 5 surface r d nd νd Medium  1 INF 0.1700 1.52426 54.3 COVER GLASS 2 INF 0.3200 1.40430 52.0 OIL 2  3 INF 0.5000 1.45852 67.8 GLASS 10  4−1.6496 4.2001 1.88298 40.8 GLASS 2  5 −3.6502 0.1200  6 −62.3111 3.22721.60298 65.4 GLASS 11  7 −8.8442 0.2000  8 26.7434 5.9486 1.49692 81.5GLASS 5  9 −7.9276 1.0000 1.63797 42.4 GLASS 6 10 132.5259 3.70731.43873 95.2 GLASS 7 11 −11.6369 0.8820 12 33.2939 1.0000 1.63797 42.4GLASS 6 13 9.1275 4.8510 1.43873 95.2 GLASS 7 14 −19.5522 0.2000 1513.6626 4.5169 1.43873 95.2 GLASS 7 16 −9.9845 1.0000 1.63797 42.4 GLASS6 17 8.8271 0.8290 18 7.1623 4.6101 1.60298 65.4 GLASS 11 19 −11.04124.4761 1.63797 42.4 GLASS 6 20 6.4981 2.5000 21 −3.7952 0.8000 1.5163064.2 GLASS 1 22 40.4882 1.8956 23 −35.9211 2.4061 1.73800 32.2 GLASS 924 −7.24980 β = 60x, NA = 1.3, field number = 22, WD = 0.32, f = 3 f(Ga) = 5.006 f (Gb) = 28.159 f (Gc) = −43.184 f (Gd) = 34.104 f (Ge) =−68.746 f (Ga) = 1.67 f (Gb) = 9.39 f (Gc) = −14.39 f (Gd) = 11.37 f(Ge) = −22.92 H1 = 7.32 H2 = 4.49 H3 = 3.9 νd (Len) = 64.2 νd (Lep) =32.2 nd (Len) = 1.51630 nd (Lep) = 1.73800 (1) H2/H1 = 0.61 (2) f (Gb)/f= 9.39 (3) νd (Len) − νd (Lep) = 32.0 (6) H3/H1 = 0.53 (9) |f (Gc)/f| =14.4

With the above-mentioned configuration, the aberration correction on thechromatic aberration and various aberrations are corrected in thewavelength range from 405 nm to 656 nm according to the presentembodiment. The aberration correction is shown in FIG. 10.

Embodiment 6

The embodiment 6 according to the present invention has theconfiguration shown in FIG. 11. That is, it is configured by, in orderfrom the object side to the image side, a positive lens group Ga, apositive lens group Gb, a lens group Gc, a lens group Gd, and a lensgroup Ge. The positive lens group Ga includes a cemented lens G1including a plano-convex lens whose plane surface faces the object sideand a meniscus lens whose concave surface faces the object side, apositive meniscus lens G2 whose concave surface faces the object side,and a positive single lens G3. The positive lens group Gb is formed by a3-cemented lens G4 including a positive lens, a negative lens, and apositive lens. The lens group Gc is formed by a cemented lens G5including a negative lens and a positive lens, and a cemented lens G6including a positive lens and a negative lens. The lens group Gd isformed by a cemented lens G7 including a positive lens and a negativelens, and shaped a meniscus lens which has the strongly concave surfacethat faces the image side. The lens group Ge is formed by a negativelens G8 including a negative lens and a positive lens.

As recognized from FIG. 11, the present embodiment shows thecharacteristic geometry of the lens group Ga on the object side.Generally, in an immersion type objective having a large NA value, thelens group of this portion shows a geometry close to a hemisphere.However, in the present embodiment, including two hemispherical lensesis one of the characteristics. With this configuration, the amount ofaberration occurring in the lens group Ga is reduced. The radius of acurvature of the image side of the cemented lens G1 is smaller than thefocal length f, and the radius of a curvature of the image side of themeniscus lens G2 is smaller than two times the focal length f. Fromthese characteristics, the ray height of and after the lens group G3 canbe reduced, thereby easily correcting a high order aberration (sphericalaberration, comatic aberration), reducing the outer diameter of theentire lens, and improving the workability of the lens.

The data according to the embodiment 1 is listed below. The objectiveaccording to the embodiment 1 is used with the immersing solution ofnd=1.51483, νd=41.0. The immersing solution is conventional oil for amicroscope objective.

TABLE 6 surface r d nd νd Medium  1 INF 0.1700 1.52426 54.3 COVER GLASS 2 INF 0.1600 1.51483 41.0 OIL 1  3 INF 0.4500 1.51630 64.2 GLASS 1  4−1.1911 2.4899 1.88298 40.8 GLASS 2  5 −2.4236 0.1500  6 −4.8347 4.73121.58909 61.1 GLASS 3  7 −5.4383 0.0996  8 INF 1.9585 1.56879 71.4 GLASS4  9 −17.9821 0.1819 10 15.6845 6.3021 1.49692 81.5 GLASS 5 11 −9.58051.0000 1.63797 42.4 GLASS 6 12 30.7512 3.7597 1.43873 95.2 GLASS 7 13−12.7994 0.2000 14 13.5205 2.7514 1.43873 95.2 GLASS 7 15 −33.24011.0000 1.63797 42.4 GLASS 6 16 6.4397 0.6500 17 7.0014 4.5120 1.4387395.2 GLASS 7 18 −11.7519 1.0000 1.63797 42.4 GLASS 6 19 −111.2141 0.556420 5.7331 6.0366 1.49692 81.5 GLASS 5 21 −6.3221 1.0000 1.61336 44.5GLASS 12 22 3.4357 2.3800 23 −3.3602 2.1379 1.51630 64.2 GLASS 1 24−16.5008 5.5869 1.80000 29.8 GLASS 14 25 −8.1656 β = 60x, NA = 1.4,field number = 22, WD = 0.16, f = 3 f (Ga) = 4.835 f (Gb) = 24.100 f(Gc) = −66.770 f (Gd) = −30.153 f (Ge) = −436.809 f (Ga) = 1.61 f (Gb) =8.03 f (Gc) = −22.26 f (Gd) = −10.05 f (Ge) = −145.60 f (G1 + G2) =5.740 H1 = 7.12 H2 = 4.47 H3 = 4.2 D = 1.33 νd (Len) = 64.2 νd (Lep) =29.8 nd (Len) = 1.51630 nd (Lep) = 1.80000 (1) H2/H1 = 0.63 (2) f (Gb)/f= 8.03 (3) νd (Len) − νd (Lep) = 34.4 (6) H3/H1 = 0.59 (7) f (G1 + G2)/f= 1.91 (8) D/f = 0.44

With the above-mentioned configuration, the aberration correction on thechromatic aberration and various aberrations are corrected in thewavelength range from 405 nm to 656 nm according to the presentembodiment. The aberration correction is shown in FIG. 12.

Embodiment 7

The embodiment 7 according to the present invention has theconfiguration shown in FIG. 13. That is, it is configured by, in orderfrom the object side to the image side, a positive lens group Ga, apositive lens group Gb, a lens group Gc, a lens group Gd, and a lensgroup Ge. The positive lens group Ga includes a cemented lens G1including a plano-convex lens whose plane surface faces the object sideand a meniscus lens whose concave surface faces the object side, apositive meniscus lens G2 whose concave surface faces the object side,and a positive single lens G3. The positive lens group Gb is formed by a3-cemented lens G4 including a positive lens, a negative lens, and apositive lens. The lens group Gc is formed by a cemented lens G5including a negative lens and a positive lens, and a cemented lens G6including a positive lens and a negative lens. The lens group Gd isformed by a cemented lens G7 including a positive lens and a negativelens, and shaped a meniscus lens which has the strongly concave surfacethat faces the image side. The lens group Ge is formed by a negativelens G8 including a negative lens and a positive lens.

As recognized from FIG. 13, the present embodiment shows thecharacteristic geometry of the lens group Ga on the object side. In thepresent embodiment, including two hemispherical lenses in the lens groupGa is one of the characteristics. With this configuration, the amount ofaberration occurring in the lens group Ga is reduced. The radius of acurvature of the image side of the cemented lens G1 is smaller than thefocal length f, and the radius of a curvature of the image side of themeniscus lens G2 is smaller than two times the focal length f. Fromthese characteristics, the ray height of and after the lens group G3 canbe reduced, thereby easily correcting a high order aberration (sphericalaberration, comatic aberration), reducing the outer diameter of theentire lens, and improving the workability of the lens.

The lens data according to the embodiment 7 is listed below. Theobjective according to the embodiment 7 is used with the immersingsolution of nd=1.51483, νd=41.0. The immersing solution is conventionaloil for a microscope objective.

TABLE 7 surface r d nd νd Medium  1 INF 0.1700 1.52426 54.3 COVER GLASS 2 INF 0.1600 1.51483 41.0 OIL 1  3 INF 0.4800 1.51630 64.2 GLASS 1  4−1.0707 1.9082 1.88298 40.8 GLASS 2  5 −2.0150 0.1500  6 −4.6997 4.04821.58909 61.1 GLASS 3  7 −4.6584 0.1000  8 INF 1.8853 1.56879 71.4 GLASS4  9 −15.6297 0.2000 10 57.7398 6.5664 1.49692 81.5 GLASS 5 11 −6.91871.0000 1.63797 42.4 GLASS 6 12 36.0738 4.4423 1.49692 81.5 GLASS 5 13−10.2050 2.1217 14 22.5473 3.9658 1.43873 95.2 GLASS 7 15 −13.23491.0990 1.63797 42.4 GLASS 6 16 25.6089 0.5967 17 10.3306 4.4472 1.4387395.2 GLASS 7 18 −10.1548 1.0000 1.63797 42.4 GLASS 6 19 122.4166 0.144020 5.0977 4.7091 1.49692 81.5 GLASS 5 21 −9.8163 1.0000 1.63797 42.4GLASS 6 22 2.9264 2.1844 23 −2.9053 1.7874 1.48745 70.3 GLASS 8 24−30.6009 5.2239 1.73800 32.2 GLASS 9 25 −7.1509 β = 60x, NA = 1.4, fieldnumber = 22, WD = 0.16, f = 3 f (Ga) = 3.973 f (Gb) = 28.967 f (Gc) =3756.617 f (Gd) = −19.235 f (Ge) = 671.695 f (Ga) = 1.32 f (Gb) = 9.66 f(Gc) = 1252.21 f (Gd) = −6.41 f (Ge) = 223.90 f (G1 + G2) = 4.5 H1 =7.01 H2 = 4.16 H3 = 4.2 D = 1.3 νd (Len) = 70.3 νd (Lep) = 32.2 nd (Len)= 1.48745 nd (Lep) = 1.73800 (1) H2/H1 = 0.59 (2) f (Gb)/f = 9.66 (3) νd(Len) − νd (Lep) = 38.1 (6) H3/H1 = 0.60 (7) f (G1 + G2)/f = 1.50 (8)D/f = 0.43

With the above-mentioned configuration, the aberration correction on thechromatic aberration and various aberrations are corrected in thewavelength range from 405 nm to 656 nm according to the presentembodiment. The process of the correction is shown in FIG. 14.

FIG. 23 shows the axial chromatic aberration from 400 nm to 700 nmaccording to the embodiments 1 through 7. For comparison with the priorart, the objectives disclosed in the Japanese Published PatentApplication No. H7-35983 and the Japanese Published Patent ApplicationNo. 2003-21786 are also described. The comparison example 1 shown inFIG. 23 is the objective disclosed in the embodiment 1 in the JapanesePublished Patent Application No. H7-35983, and the comparison example 2shown in FIG. 23 is the objective disclosed in the embodiment 2 of theJapanese Published Patent Application No. H7-35983. The comparisonexample 3 is the objective disclosed in the embodiment 4 of the JapanesePublished Patent Application No. 2003-21786. The sectional view of thelens in the comparison example 1 is shown in FIG. 15, and FIG. 16 showsthe aberration. FIG. 17 shows the sectional view of the lens of thecomparison example 2, and FIG. 18 shows the aberration. FIG. 19 showsthe sectional view of the lens of the comparison example 3. FIG. 20shows the aberration.

FIG. 23 shows the focal point on the axis on the object plane at eachwavelength normalized on the e-line (546.07 nm). The horizontal axisindicates the wavelength (nm), and the vertical axis indicates the gapof the focal point at each wavelength when the e-line (546.07 nm) isdefined as the zero.

As shown in FIG. 23, the axial chromatic aberration of the objectiveaccording to the embodiments of the present invention is appropriatelycorrected so that the gap of the focal point at each wavelength iswithin 0.1 μm (0.0001 mm) in 400 nm through 700 nm. This means theobjective according to an embodiment of the present inventioneffectively works on a multi-wavelength observation.

In addition, as clearly shown in comparing the aberration (FIGS. 2, 4,6, 8, 10, 12, and 14) according to each embodiment with the aberration(FIGS. 16, 18, and 20) of each comparison example, the embodiments ofthe present invention have characteristics in the method of correctingthe spherical aberration. As shown in FIG. 24B, the objective accordingto the present embodiment corrects the spherical aberration equally onall NA at each wavelength. On the other hand, in the comparison exampleas the prior art, as shown in FIG. 24A, there are positive and negativeportions of the spherical aberration with respect to the NA, whichoffsets the focal points for the respective wavelengths, therebysubstantially equally correcting the focal point for each wavelength.

When the aberration correction as in the above-mentioned prior art isperformed, there occurs a serious effect when the NA of the objective isused with bias. For example, the effect clearly comes out when the NA ofthe objective is narrowed and the laser beam having a Gaussiandistribution is input to the objective.

FIG. 25 is an explanatory view of the event. As shown in FIG. 25, with acommon and conventional objective, the chromatic aberration of a rayfrom the position of a larger NA is corrected such that the ray can forman image at substantially the same position regardless of thewavelength, but a ray at the position of a smaller NA forms an image ata different position for each wavelength. The direct cause of the eventis the method of correcting the spherical aberration as shown in FIG.24A.

On the other hand, with the objective according to an embodiment of thepresent invention, as shown in FIG. 24B, since the spherical aberrationis equally corrected over the entire NA for each wavelength, thereoccurs little influence although the NA of the objective is used withbias. That is, the objective according to the present invention ishighly appropriate for the laser scanning microscope used by a Gaussianbeam incident to the objective.

FIG. 23 shows the chromatic aberration of magnification of theembodiments 1 through 7 and comparison example, which is the chromaticaberration of magnification at field number 9 normalized at thewavelength 488 nm. The horizontal axis indicates the wavelength (nm),and the vertical axis indicates the gap of the focal point at eachwavelength. The field number 9 corresponds to 0.075 mm in the objectiveplane since the objectives magnifies object 60 times.

As shown in FIG. 26, the chromatic aberration of magnification of theobjective according to the embodiments of the present invention isappropriately corrected so that the gap of the focal point at eachwavelength is within 0.3 μm (0.0003 mm). Described below are theembodiments of the present invention from the point of the fluorescentobservation apparatus.

The fluorescent observation apparatus as one of the embodiments of thepresent invention includes: a light source; a beam splitting deviceselectively reflecting a ray from the light source; an objective forilluminating or observing a specimen; a stage for fixing a specimen; awavelength selection device for selecting a desired wavelength rangebased on rays that pass through the beam splitting device; and adetector for detecting a ray that passes through the wavelengthselection device. With the configuration, assume that λ1 is the longestwavelength and λ2 is the shortest wavelength in illuminating thespecimen using two or more wavelengths, and Δ1 and Δ2 are the focalpoint of λ1 and λ2 respectively on the optical axis of the objective.Based on the assumption, the following conditions are satisfied.λ1−λ2≧180 nm  (11)|Δ1−Δ2|<0.2 μm  (12)

If the expressions (11) and (12) can be satisfied, then the chromaticaberration can be ignored although the specimen is dyed with a pluralityof fluorescent tags.

It is furthermore desired to satisfy the following conditionalexpression.λ2≦442 nm  (13)

Recently used is a photoactivation fluorescent tag whose characteristicis changed by a stimulating ray on the shorter wavelength side. Forexample, with photoconvertion fluorescent protein such as Kaede, PA-GFP,etc., a wavelength of 405 nm is commonly used. In addition, since 442 nmis used in the observation of fluorescent resonant energy transfer(FRET) of CFP-YFP, the observations of the photoactivation andfluorescent resonant energy transfer (FRET) can be performed if theexpression (13) is satisfied.

It is furthermore desired to satisfy the following condition.|δ1−δ2|≦0.3 μm  (14)

where δ1 and δ2 indicate the chromatic aberration of the magnificationof each wavelength at the field number 9 of λ1 and λ2 respectively.

At this time, the objective is configured by, in order from the objectside to the image side, a positive lens group Ga, a positive lens groupGb, a lens group Gc, a lens group Gd, and a lens group Ge. The positivelens group Ga includes a cemented lens obtained by cementing aplano-convex lens whose plane surface faces the object side to ameniscus lens whose concave surface faces the object side, and apositive single lens. The positive lens group Gb is formed by a cementedlens. The lens group Gc includes one or more cemented lens. The lensgroup Gd includes a meniscus lens having a strongly concave surface thatfaces the image side. The lens group Ge includes a negative lens havinga strongly concave surface that faces the object side. It can beconsidered that the lens group Gb is a cemented triplet including apositive lens, a negative lens, and a positive lens. Using the objectivewith the configuration is suitable for the fluorescent observation.

Furthermore, it is desired that the objective satisfies the followingconditions.0.5≦H2/H1≦0.75  (1)7.8≦f(Gb)/f≦20  (2)

In addition, it is desired that the objective compensate the chromaticaberration by itself. In this case, the flexibility of selecting theunits to be combined (image optics system, illumination optics system)is enhanced.

The fluorescent observation apparatus as one of the embodiments of thepresent invention includes: an image storage device for storing at leasttwo images formed from a detection result by the detector when thespecimen is illuminated using the excitation wavelengths λ1 and λ2; andan image analysis device for calculating the overlap, the difference, orthe ratio of the images. With the configuration, it is appropriate tocalculate the overlap, the difference, or the ratio of a plurality ofimages.

the apparatus also includes an image storage device for storing an imageexcited by the excitation wavelengths λ1 and λ2, and an image analysisdevice for calculating the overlap, the difference, or the ratio of theimages. With the configuration, it is easy to analyze the molecules inthe specimen dyed with a fluorescent tag by calculating the overlap, thedifference, or the ratio of a plurality of images. Recently, multipledata of a plurality of fluorescent wavelengths is obtained, for example,the FRET for analysis of molecules. In this case, a chromatic aberrationcorrected microscope objective is appropriate.

Furthermore, with the fluorescent observation apparatus as one of theembodiments of the present invention, the stage or the objective can bemoved in the direction of the optical axis. At least two images formedas the detection result by the detector when the specimen is illuminatedwith the different excitation wavelengths λ1 and λ2. The above procedureis repeated while the intervals of the specimen and the objective arechanged step by step, and the sectional images of the specimen with thedifferent excitation wavelengths λ1 and λ2. Finally, multicolored threedimensional image of the specimen is formed, overlapping the threedimensional images with wavelengths λ1 and λ2.

The present chromatic aberration corrected system is specificallyeffective in generating a three-dimensional image of a specimen. Ifthere is axial chromatic aberration, the focal point in the direction ofthe optical axis with λ1 and λ2 is shifted in the direction of theoptical axis. Therefore, when the images of λ1 and λ2 are overlapped, ithas conventionally been necessary to overlap the images having differentsectional positions in the axial chromatic aberration of the objective.In the present system, the axial chromatic aberration is so small thatthe axial chromatic aberration can be ignored. As a result, a correctthree dimensional image can be generated.

The fluorescent observation apparatus as one of the embodiments of thepresent invention include: a scanning device scanning a beam spot on thefocal plane; and a confocal detection device projecting the beam spot.

The fluorescent observation apparatus as one of the embodiments of thepresent invention is appropriate for observation on the threedimensional image of the specimen since the axial chromatic aberrationis corrected.

The fluorescent observation apparatus as one of the embodiments of thepresent invention include: a plurality of laser light sources areprovided to combine lasers of plural wavelengths on the same opticalpath, and lead the combined lasers into the fluorescent observationapparatus by one optical fiber.

When laser lights are led into a microscope, it is usual to employ fiberoptics which has high flexibility. In conventional art, differentoptical fiber are used in case that the desired lasers have differencein the wavelength. In this configuration, adjusting the subsequentlyarranged collimating lenses can correct the chromatic aberration, but itis hard to adjust the collimating lenses for each. On the other hand,since the fluorescent observation apparatus as one of the embodiments ofthe present invention is corrected in the chromatic aberration, thisfluorescent observation apparatus can employ a broadband optical fiber,which is developed recently.

The fluorescent observation apparatus as one of the embodiments of thepresent invention is appropriate for observing a specimen indicated by aplurality of fluorescent tags, the position information about themolecules in the specimen provided with two or more indicators isanalyzed by superposing and outputting images obtained by the excitationwavelengths of λ1 and λ2.

In case of observing fluorescent resonant energy transfer orphotoconversion, it is necessary to compute the ratio or composite imageof the fluorescence of the different wavelength. In this observation, ithappens to obtain inaccurate data caused by the chromatic aberration.The fluorescent observation apparatus as one of the embodiments of thepresent invention is appropriate for observing the information where themolecules are in the specimen.

The fluorescent observation apparatus as one of the embodiments of thepresent invention is appropriate for obtaining a plurality of imagesformed from the detection result by the detector when the specimen isilluminated by the excitation wavelengths of λ1 and λ2 are acquired withthe lapse of time, and the fluorescent resonant energy transfer betweenthe molecules in the specimen is observed by obtaining a plurality ofratios of images by λ1 and λ2 with the lapse of time when the images areacquired at the same time.

In conventional art, it is often necessary to composite the images ofdifferent sections on the optical axis intentionally because of thechromatic aberration. In this technique, the gap between the images ofthe different wavelength can be compensated when the specimen is stay.However it is necessary to observe the timely accurate data in entirewavelengths in case of observing the molecular migration in a livespecimen. The fluorescent observation apparatus as one of theembodiments of the present invention is appropriate for the observationswhich trace rapid molecular migrations.

The fluorescent observation apparatus as one of the embodiments of thepresent invention is appropriate for obtaining a specimen image havingtwo or more fluorescent tags is acquired and brightness informationabout the obtained image is expressed and plotted in a two-dimensionalarray for each pixel, thereby estimating localization of each molecule.

Colocalization method is to obtain images of a specimen with at leasttwo fluorescent tags and to plot the brightness information of theimages and to estimate localization of molecules in the specimen. Thefluorescent observation apparatus as one of the embodiments of thepresent invention is appropriate for this observation.

The fluorescent observation apparatus as one of the embodiments of thepresent invention is appropriate for outputting an image of specimenformed from a detection result of the detector when a excitationilluminate the specimen, specifying an area of a part of the image ofthe specimen, photobleaching fluorescence tags in the specified area,recording the time change of the fluorescent image in the specified areaor the outside, and detecting the molecular dispersion in the specimen.

Fluorescent recovery after photobleaching (FRAP) is to output an imageof specimen formed from a detection result of the detector when aexcitation illuminate the specimen, to specify an area of a part of theimage of the specimen, to photobleach fluorescence tags in the specifiedarea, to record the time change of the fluorescent image in thespecified area or the outside, and to detect the molecular dispersion inthe specimen. The fluorescent observation apparatus as one of theembodiments of the present invention is appropriate for thisobservation, because the chromatic aberration is corrected.

The fluorescent observation apparatus as one of the embodiments of thepresent invention is appropriate for outputting an image of specimenformed from a detection result of the detector when a excitationilluminate the specimen, specifying an area of a part of the image ofthe specimen, photoactivating fluorescence tags in the specified area,recording the time change of the fluorescent image in the specified areaor the outside, and for analyzing the molecules in the specimen.

The fluorescent observation apparatus as one of the embodiments of thepresent invention is appropriate for the observation with Kaede orPA-GFP etc., which are photoactivated by the light of wavelength 405 nm,because the chromatic aberration is corrected in the range betweenwavelength 405 nm and 653 nm.

The fluorescent observation apparatus as one of the embodiments of thepresent invention is appropriate for illuminating a specimen with aplurality of fluorescent tags by a plurality of excitement lightsimultaneously, detecting the fluorescent lights which are separated,for recording the time change of the fluorescent lights, and computingthe velocity of the molecules in the specimen.

There are methods called “FCS” and “FCCS”, which is to record the timechange of fluorescence and calculate the correlation function. When aplurality of fluorescent wavelengths are detected using a plurality ofexcitation wavelengths, it is confirmed that molecules are coupled. Inthis case, since it is necessary that there is no gap in excitationwavelengths on the condensing spot to a specimen, the fluorescentobservation apparatus as one of the embodiments of the present inventionis appropriate.

FIG. 27 shows the configuration of a conventional laser scanningmicroscope. When, for example, an argon ion laser (wavelength of 488nm), a He—Ne green laser (wavelength of 543 nm), and a He—Ne laser(wavelength of 633 nm) are used as a visible laser 2, the lasers arecombined by a dichroic mirror 4 on the same optical path, and condensedinto a single mode fiber 8 by a converging lens 6. Since the single modefiber has an NA of about 0.1, and the diameter of the core of severalμm, a strict adjustment is required to put the laser beams into thefiber. Therefore, a fiber coupling mechanism 7 is provided for adjustingthe tilt and the horizontal shift of the laser beams with respect to thefiber.

At this time, when a laser having the wavelength of 405 nm etc. isfurther used, another fiber is often used as a short wavelength laser 1,introduced to a laser scanning microscope, and combined by a dichroicmirror 12 at a later stage in the conventional technology because thechromatic aberration caused by the objective 18 is adjusted by acollimator lens 10 after the fiber 8. The adjustment is required to usea special method when a laser having the wavelength of 405 nm etc. isused because the wavelength range in which the chromatic aberration iscorrected is not sufficient with the conventional objective.

However, since each objective has different chromatic aberrations, it isvery difficult to use this adjusting method for all objectives. Sincethe diameter of an optical fiber is very small and is only severalmicrons, it is necessary to adjust a fiber tilt alignment mechanism 9for strict parallelism and tilt of the fiber when it is adapted to amicroscope. Therefore, the conventional laser scanning microscoperequires complicated adjustments.

FIG. 28 shows an embodiment of the laser scanning microscope accordingto the present invention. Since the chromatic aberration of theobjective according to the present embodiment is corrected in thewavelength range from 406 nm to 656 nm, the laser on the shortwavelength laser and the visible laser can be handled similarly.Therefore, the single mode fiber 8 is used. The fiber can be a broadbandfiber having a broad wavelength range available.

The beams from the fiber 8 are parallel beams through the collimatorlens 10, and the visible laser and the laser on the short wavelengthlaser are combined into the same optical path by a half mirror or adichroic mirror, and reflected by a dichroic mirror 13. The laser beamreflected by the dichroic mirror is scanned by a galvano-mirror 14,relayed by a pupil projection lens 15 and a tube lens 16, and led intoan objective. The fluorescence from a specimen passes through an inverseoptical path, passes through a dichroic mirror 13, and is condensed to apinhole 19 for a confocal effect by a tube lens 20. After distributingthe fluorescence that has passed through the pinhole 19 using a dichroicmirror 21, fluorescence in a desired wavelength range is detected by aphotodetector 24 using a barrier filter 23. The reference numerals 3, 5,11, 17, and 22 designate a mirror for bending an optical path.

It is further desired that the dichroic mirror 13 is a multibanddichroic mirror capable of reflecting lasers of a plurality ofwavelengths without switching the dichroic mirror 13. When a dichroicmirror has a wedge, the condensing position is a little shifted on thesample surface by a switch of a dichroic mirror. However, using amultiband dichroic mirror, one dichroic mirror can process a pluralityof lasers. Therefore, no switching is required, and no shift of thecondensing position is detected on the sample surface by the wedge ofthe dichroic mirror. When a plurality of objectives is used, theconventional technology cannot correctly apply the chromatic aberrationto a plurality of objectives. However, using the low chromaticaberration objective as one of the embodiments of the present invention,and with the configuration of the fluorescent observation apparatus, anobservation has little error caused by chromatic aberration.

The objective according to the present embodiment compensates thechromatic aberration by itself. Therefore, it is advantageous when alaser beam is introduced between the objective 18 and the tube lens 16.When the chromatic aberration correction is performed by compensatingfor the tube lens 16 and the objective 18, there occurs a shift in afocal point is a laser etc. is introduced between the objective 18 andthe tube lens 16. On the other hand, there is no shift occurring in thefocal point according to the present embodiment in which the chromaticaberration correction is performed using only the objective. Therefore,it is advantageous when an observation is performed by a laser scanningmicroscope through a stimulus by a laser beam.

FIG. 28 shows the configuration of a stimulating laser 33 provided byarranging a dichroic mirror 32 between the objective 18 and the 17. Inthis example, a movable mirror 34 is provided for adjusting theirradiation position of the stimulating laser 33.

FIG. 29 shows an example of the configuration of the appearance of thelaser scanning mirror focal microscope as an embodiment of the presentinvention. A specimen 25 is placed on a stage 26, and an objective 18 isattached to the revolver of a microscope body 28. A plurality ofobjectives can be attached to the revolver. The objective has anobjective lift mechanism 27 that travels in the direction of the opticalaxis, and a plurality of sectional images of the specimen 25 in thedirection of the optical axis can be obtained. To obtain athree-dimensional image of a specimen, a confocal scanner 29 is attachedto a microscope body 28, and a galvano-scanner, a pin hole, etc. arearranged inside. Although not shown in the attached drawings, a laserlight source is connected to the confocal scanner 29 through a fiber asshown in FIG. 27. A signal obtained by the confocal scanner 29 istransmitted to an image processing device 31, and displayed on an imagedisplay device 30. To generate a three-dimensional image of a specimen,the objective is moved in the direction of the optical axis, and afluorescent image of a plurality of wavelengths is acquired. Eachsectional image is stored in the image processing device 31, and athree-dimensional image of a specimen is displayed on the image displaydevice 30.

Since the chromatic aberration is sufficiently corrected according tothe present invention, there is no shift in the direction of the opticalaxis on a plurality of fluorescent images. Therefore, the sectionalimages of the specimen stored in the image processing device 31 andhaving a plurality of fluorescent wavelengths in the direction of theoptical axis are superposed and displayed, correct three-dimensionalimages of the specimen can be obtained only by superposing the samesectional images different in wavelengths. Furthermore, when a fastmovement of a molecule in a specimen is viewed, the configuration of thepresent embodiment requires no driving of an objective in the directionof the optical axis for chromatic aberration correction when a pluralityof fluorescent images are superposed. Therefore, a fast movement of amolecule in a specimen can be observed.

Furthermore, the present invention is also effective in the fluorescentmicroscope and the fluorescent observation apparatus other than theconfocal microscope. For example, there is an observation method ofchecking how a protein moves from the cell membrane by indicating theprotein in a cell by fluorescent tags having different wavelengths. Inthis case, it is necessary to bring the cell membrane itself into thefocus of a microscope. When there is axial chromatic aberration in theobjective, the lens may go out of focus by different fluorescent tags,thereby observing a position shifted from the cell membrane by adifferent fluorescent wavelength. Also in this case, the same plane of asample can be constantly observed using the objective as one of theembodiments of the present invention when fluorescent tags havingdifferent fluorescent wavelengths are used, thereby obtaining correctdata.

FIGS. 30A, 30B, and 30C show an example of observing the fluorescentresonant energy transfer (FRET) as an application of the fluorescentobservation apparatus according to the embodiments of the presentinvention. In this observation, the density of the calcium ion in thecell is measured using, for example, the fluorescent probe of Cameleon.The Cameleon has the structure of two types of fluorescent protein ofCFP and YFP coupled by the protein such as calmodulin-M etc. When thedensity of the calcium ion in the cell is low, only the fluorescencehaving the wavelength of 485 nm is emitted from the CFP when the excitedlight of 442 nm is emitted. However, if the density of the calcium ionbecomes high, energy is transferred from the CFP to the YFP, and thefluorescence having the wavelength of 530 nm is observed as thefluorescence from the YFP. Based on the phenomenon, the ratio of theintensity of the fluorescence between the CFP and the YFP is measured,thereby measuring the density of the calcium ion.

A fluorescent microscope or a laser scanning microscope acquires animage by two wavelengths of 485 nm and 530 nm. For example, FIG. 30Ashows an image of 485 nm, and FIG. 30B shows image of 530 nm. FIG. 30Cshows the image for which the ratio of the brightness is obtained foreach pixel of the images FIGS. 30A and 30B. The image FIG. 30C obtainedin this operation has the information about the density of the calciumion in the cell. As clearly shown in the above-mentioned procedure, itis necessary in this observation method to correctly associate the imageacquired with a different wavelength with each pixel. The fluorescentobservation apparatus according to the embodiments of the presentinvention is appropriate for the observation.

Described above is the fluorescent resonant energy transfer(FRET), butin the observation method of taking multiple values of fluorescence inmeasuring the calcium such as fura 2 etc., multiple values can beconstantly taken on the same plane of a sample plane according to one ofthe embodiments of the present invention, thereby realizing a correctmeasurement.

1. An immersion type microscope objective comprising, in order from an object side to an image side, a positive lens group Ga, a positive lens group Gb, a lens group Gc, a lens group Gd, and a lens group Ge, wherein: the positive lens group Ga includes a cemented lens obtained by cementing a plano-convex lens whose plane surface faces the object side and a meniscus lens whose concave surface faces the object side, and a positive single lens; the positive lens group Gb is formed by a cemented lens; the lens group Gc includes at least one cemented lens; the lens group Gd includes a meniscus lens having a strongly concave surface that faces the image side; the lens group Ge includes a negative lens Len having a strongly concave surface that faces the object side, and a positive lens Lep; wherein the following conditions are satisfied: 0.5≦H2/H1≦0.75, 0.5≦H3/H1≦0.65, and 7.8≦|f(Gb)/f|≦20, where H1 indicates a ray height of a marginal ray emergent from the lens group Gb, H2 indicates a ray height of a marginal ray incident to the lens group Gd, H3 indicates a ray height of a marginal ray emergent from the lens group Ge, f indicates a focal length of the entire objective, and f(Gb) indicates a focal length of the lens group Gb; and wherein the following condition is satisfied: 45≧νd(Len)−νd(Lep)≧30, where νd(Len) is an Abbe number of glass of the negative lens Len, and νd(Lep) is an Abbe number of glass of the positive lens Lep.
 2. The immersion type microscope objective according to claim 1, wherein the glass of the positive lens Lep satisfies the following conditions: 1.65≦nd(Lep)≦1.8, 25≦νd(Lep)≦41, wherein nd (Lep) is a d-line refractive index of the positive lens Lep.
 3. The immersion type microscope objective according to claim 1, wherein the lens group Gb is a cemented triplet comprising a positive lens, a negative lens, and a positive lens.
 4. An immersion type microscope objective comprising, in order from an object side to an image side, a positive lens group Ga, a positive lens group Gb, a lens group Gc, a lens group Gd, and a lens group Ge, wherein: the positive lens group Ga comprises: (i) a cemented lens G1 obtained by cementing a plano-convex lens whose plane surface faces the object side and a meniscus lens whose concave surface faces the object side, (ii) a positive meniscus lens G2 whose concave surface faces the object side, and (iii) a positive single lens G3; the positive lens group Gb is formed by a cemented lens; the lens group Gc includes at least one cemented lens; the lens group Gd includes a meniscus lens having a strongly concave surface that faces the image side; the lens group Ge includes a negative lens having a strongly concave surface that faces the object side; and wherein the following conditions are satisfied: 0.5≦H2/H1≦0.75, 7.8≦|f(Gb)/f|≦20, 1≦f(G1+G2)/f≦2, and D/f≦0.6, where f(G1+G2) is a composite focal length of the cemented lens G1 and the meniscus lens G2, f is a focal length of the entire objective, D is a length of a marginal ray between the cemented lens G1 and the meniscus lens G2, H1 indicates a ray height of a marginal ray emergent from the lens group Gb, H2 indicates a ray height of a marginal ray incident to the lens group Gd, and f(Gb) indicates a focal length of the lens group Gb.
 5. An immersion type microscope objective comprising, in order from an object side to an image side, a positive lens group Ga, a positive lens group Gb, a lens group Gc, a lens group Gd, and a lens group Ge, wherein: the positive lens group Ga includes a cemented lens obtained by cementing a plano-convex lens whose plane surface faces the object side and a meniscus lens whose concave surface faces the object side, and a positive single lens; the positive lens group Gb is formed by a cemented lens; the lens group Gc includes at least one cemented lens; the lens group Gd includes a meniscus lens having a strongly concave surface that faces the image side; the lens group Ge includes a negative lens having a strongly concave surface that faces the object side; wherein the following conditions are satisfied: 0.5≦H2/H1≦0.75, and 7.8≦|f(Gb)/f|≦20, where H1 indicates a ray height of a marginal ray emergent from the lens group Gb, H2 indicates a ray height of a marginal ray incident to the lens group Gd, f indicates a focal length of the entire objective, and f(Gb) indicates a focal length of the lens group Gb; and wherein the lens group Gc is movable in a direction of an optical axis, and the following condition is satisfied: |f(Gc)/f|≦50 where f is the focal length of the entire objective, and f(Gc) is a focal length of the lens group Gc.
 6. The immersion type microscope objective according to claim 5, wherein the following condition is further satisfied: 10≦|f(Gc)/f|≦20.
 7. The immersion type microscope objective comprising, in order from an object side to an image side, a positive lens group Ga, a positive lens group Gb, a lens group Gc, a lens group Gd, and a lens group Ge, wherein: the positive lens group Ga includes a cemented lens obtained by cementing a plano-convex lens whose plane surface faces the object side and a meniscus lens whose concave surface faces the object side, and a positive single lens; the positive lens group Gb is formed by a cemented lens; the lens group Gc includes at least one cemented lens; the lens group Gd includes a meniscus lens having a strongly concave surface that faces the image side; the lens group Ge includes a negative lens having a strongly concave surface that faces the object side; and wherein the following conditions are satisfied: 0.5≦H2/H1≦0.75, 7.8≦|f(Gb)/f|≦20, λ1+λ2≧180 nm, and |Δ1−Δ2|<0.2 μm, where H1 indicates a ray height of a marginal ray emergent from the lens group Gb, H2 indicates a ray height of a marginal ray incident to the lens group Gd, f indicates a focal length of the entire objective, f(Gb) indicates a focal length of the lens group Gb, λ1 and λ2 are two wavelengths of two rays that pass through the immersion type microscope objective, and Δ1 and Δ2 are focal points on an optical axis of the objective with respect to λ1 and λ2 respectively.
 8. The immersion type microscope objective according to claim 7, wherein the following condition is satisfied: λ2≦442 nm.
 9. The immersion type microscope objective according to claim 7, wherein the following condition is satisfied, |δ1−δ2|≦0.3 μm where δ1 and δ2 indicate chromatic aberration of magnification of μ1 and μ2 respectively at field number
 9. 